1. Field of the Invention
The present invention relates to a flow cytometry apparatus, and more particularly concerns a flow cytometry apparatus for detemining characteristics of cells or the like which provides for spatial wavelength separation of spectrally rich light for improved excitation energy in a flow cytometry apparatus.
2. Description of the Prior Art
Flow cytometry apparatuses rely upon the flow of cells or other particles in a liquid flow stream in order to determine one or more characteristics of the cells under investigation. For example, a liquid sample containing cells is directed through the flow cytometry apparatus in a rapidly moving liquid stream so that each cell passes serially, and substantially one at a time, through a sensing region. Cell volume may be determined by changes in electrical impedance as each cell passes through the sensing region. Similarly, if an incident beam of light is directed at the sensing region, the passing cells scatter such light as they pass therethrough. This scattered light has served as a function of cell shape, index of refraction, opacity, roughness and the like. Further, fluorescence emitted by labeled cells which have been excited as a result of passing through the excitation energy of the incident light beam is detectable for identification of specifically labeled cells. Not only is cell analysis performed on the flow cytometry apparatuses, but sorting of cells may also be achieved. Lasers have been used as the source of the incident beam of illumination in flow cytometry apparatuses, as well as sources of incoherent or non-collimated light, such as mercury or xenon arc lamps, which typically provide spectrally rich light. Such apparatuses which include incoherent light sources have been described in copending patent applications, Ser. No. 276,738, filed on June 24, 1981, in the U.S. Patent and Trademark Office, and Ser. No. 482,346, filed in the U.S. Patent and Trademark Office on Apr. 5, 1983, both applications having a common assignee herewith, and also in U.S. Pat. No. 4,348,107. One flow cytometry apparatus known and sold as the FACS.TM. Analyzer, FACS Systems, Becton, Dickinson and Company, Sunnyvale, California, presently relies upon a mercury arc lamp as the excitation source of incoherent light for illuminating the stream of particles or cells flowing therethrough.
Excitation energy from a mercury or xenon arc lamp, or other incoherent light sources, is typically bright and spectrally rich. Accordingly, these arc lamps are desirable sources of excitation energy since the spectrally rich light is composed of a group of very intense lines (wavelengths at which the energy is much higher than at other wavelengths). Ordinarily, these energy lines are selected by using optical filters which either absorb or reflect the light of the undesirable wavelengths. By utilization of this filtering technique, the intense energy lines of light are permitted to intercept the flowing stream of particles or cells in the flow cytometry apparatus. If the cells have been labeled with a fluorochrome, the intense energy lines serve as the excitation energy to cause the fluorescently labeled cells to emit fluorescence. While the known filters are efficient in removing undesirable wavelengths of light in this type of illumination scheme, the filters usually attenuate the desired wavelengths of light by up to 70%. Moreover, the filters need to be changed each time a different wavelength is selected, for example, as the excitation energy for the fluorescently labeled cells.
Accordingly, improvements are still being sought for flow cytometry apparatuses which rely upon an incoherent light source, such as mercury or xenon arc lamps which provide spectrally rich light for illumination of the particles flowing in the liquid flow stream. Improvements are needed particularly to selectively choose those desired wavelengths or lines relied upon for excitation energy for the fluorescently labeled cells. It is to such improvement that the present invention is directed.